Abstract

Acquired resistance to HER2 kinase inhibitors, such as lapatinib, presents an unmet clinical need among the 20% of breast cancers with HER2 amplification. To identify mechanisms of lapatinib resistance, we generated a lapatinib resistance model by extended exposure of the AU565 HER2-amplified breast cancer cell line to lapatinib (AU565 LapR cell line), which was highly resistant to lapatinib compared to AU565 parental cells. AU565 LapR cells maintained mTORC1 activation (indicated by p-P70 S6K and p-S6) after lapatinib treatment, whereas mTORC1 was completely inhibited in lapatinib-treated parental cells. Moreover, HER2 was inhibited equally well by lapatinib in parental and LapR cells, suggesting that mTORC1 activation in LapR cells was HER2-independent. Interestingly, mTORC1 activation in LapR cells was PI3K/Akt-independent, as p-Akt was completely inhibited by lapatinib in parental and LapR cells, while mTORC1 activation was maintained in LapR cells only. LapR cells were sensitive to mTORC1 and mTORC1/2 inhibition, but not to PI3K inhibition. Knockdown of the GTPase Rheb, which is known to promote mTORC1 activation, abolished mTORC1 activity in lapatinib-treated LapR cells, indicating that Rheb was required for mTOR activation in LapR cells. Surprisingly, several known mediators of Rheb/mTOR activation, such as TSC1/2, AMPK, the β-catenin pathway, and PRAS40, were not altered in their activation status in LapR cells compared with parental cells. This suggests that a potentially novel mechanism of mTOR activation may promote lapatinib resistance. These findings suggest the use of mTOR inhibitors in lapatinib-resistant breast cancers, and also may lead to novel basic mechanistic insights into mTOR signaling, which is commonly dysregulated in cancer.